The resulting mutant mice showed neuronal hypertrophy, and an increased number of dopaminergic neurons and fibers in the ventral mesencephalon. Interestingly, quantitative microdialysis studies in Pten KO mice revealed no alterations in basal DA extracellular levels or evoked DA release in the dorsal striatum, despite a significant increase in total DA tissue levels. Striatal dopamine receptor D1 (DRD1) and prodynorphin (PDyn) mRNA levels were significantly elevated in KO animals, suggesting an enhancement in neuronal activity associated with the striatonigral projection pathway, while dopamine receptor D2 (DRD2) and preproenkephalin (PPE) mRNA levels remained unchanged. In addition, PTEN inactivation protected DA neurons and significantly enhanced DA-dependent behavioral functions in KO mice after a progressive 6OHDA lesion. These results provide further evidence about the role of PTEN in the brain and suggest that manipulation of the PTEN/Akt signaling pathway during development may alter the basal state of dopaminergic neurotransmission and could provide a therapeutic strategy for the treatment of Parkinsons disease, and other neurodegenerative disorders. PTEN deletion or Akt/PKB activation in dopamine neurons of the ventral midbrain results in remarkable hypertrophy of the substantia nigra and VTA. Our initial characterization of a DAT-PTEN KO strain has provided a clear definition of some of the neuroadaptations in the mesolimbic and nigrostriatal systems, and clearly show dopamine neurotransmission is permanently altered in PTEN KO mice. However, while DAT-PTEN KO animals are viable and appear behaviorally competent, an in depth study of behavioral parameters will clarify if the lack of PTEN interferes with essential functions related to the dopamine system in young, adult and aged animals. Studies performed over the past few years have clearly shown that phenotypes caused by specific genetic modifications are strongly influenced by genes unlinked to the target locus. This problem is exacerbated through the use of Cre-LoxP models as two strains, often containing their own (obscure) genetic backgrounds, are crossed through very specific breeding schedules to generate control and experimental animals. Clearly, it becomes important to avoid the use of a mixed genetic background so complex as to preclude any reasonable use of controls and prevent replication by other investigators. To perform complex behavioral studies, we will use a c57bl/6 congenic DAT-PTEN KO mouse line, generated in our lab. A new mechanism in the brain of rats that may mediate the rewarding and reinforcing properties of drugs of abuse has recently been discovered. This mechanism involves the physical interaction between two proteins in midbrain dopamine neurons, the tumor suppressor PTEN and the brain specific receptor for serotonin, the 5-HT2c receptor (5-HT2cR). Blocking the interaction of PTEN with 5-HT2cR prevents the development of conditioned place preference to nicotine and THC, the active component in marijuana. In addition, PTEN has been shown to physically interact with the NR1 subunit of NMDAR in hippocampus, and PTEN downregulation decreases NMDAR surface expression. These studies suggest PTEN in dopamine neurons may directly modulate functions intimately linked to the development of addiction, and dopamine mediated cognition, such as responses to reward and motivation. We will use a congenic PTEN-KO line to analyze in detail the behavioral profile of KO animals in relationship to drug abuse, overall locomotor performance, as well as other dopamine related cognitive functions. We have shown Pten deletion in differentiated DA neurons causes a significant increase in the number and size of surviving neurons in both the mesolimbic and nigrostriatal projecting pathways (see above). Because at the time of Pten deletion DA neurons have already completed mitosis and phenotypic determination, it is unlikely the reported increase in DA neurons is due to an increase in newly formed neurons. It is thus likely PTEN ablation preserves DA neurons that normally would undergo apoptosis due to the lack of target support, by repressing the initiation of apoptotic pathways. We are now intrigued about several aspects induced by PTEN deletion in dopamine cells: Do all dopamine neurons preserved in DAT-PTEN KO animals project to target areas and form functional connections? Can exposure to an enriched environment enhance dopaminergic function in young and aged KO animals? Do the mesolimbic and nigrostriatal dopamine projections remain functional into aging? Is the PTENless aging dopaminergic system more or less resistant to neurotoxic insults applied during different stages of the mouse life span? Are PTENless dopamine neurons prone to form tumors? Obviously, the tremendous adaptations observed in the PTENless dopaminergic system during development, may pose problems for interpretation of results; however, as previous studies have shown Akt/PKB activation in the adult dopaminergic system can also result in remarkable hypertrophy and plasticity of the nigra and VTA, the results obtained in this study may provide valuable insights into how PTEN ablation changes dopamine function at the molecular and behavioral levels, and the long-term consequences of such adaptations. We believe these studies are important, as manipulations of the PTEN pathway are being considered as a possible venue for therapeutic strategies involving the brain.